The mechanisms by which the immune system achieves constant T cell numbers throughout life, thereby controlling autoaggressive cell expansions, are to date not completely understood. Here, we show that the CD25+ subpopulation of naturally activated (CD45RBlow) CD4 T cells, but not CD25− CD45RBlow CD4 T cells, inhibits the accumulation of cotransferred CD45RBhigh CD4 T cells in lymphocyte-deficient mice. However, both CD25+ and CD25− CD45RBlow CD4 T cell subpopulations contain regulatory cells, since they can prevent naive CD4 T cell-induced wasting disease. In the absence of a correlation between disease and the number of recovered CD4+ cells, we conclude that expansion control and disease prevention are largely independent processes. CD25+ CD45RBlow CD4 T cells from IL-10-deficient mice do not protect from disease. They accumulate to a higher cell number and cannot prevent the expansion of CD45RBhigh CD4 T cells upon transfer compared with their wild-type counterparts. Although CD25+ CD45RBlow CD4 T cells are capable of expanding when transferred in vivo, they reach a homeostatic equilibrium at lower cell numbers than CD25− CD45RBlow or CD45RBhigh CD4 T cells. We conclude that CD25+ CD45RBlow CD4 T cells from nonmanipulated mice control the number of peripheral CD4 T cells through a mechanism involving the production of IL-10 by regulatory T cells.

The kinetics of postnatal intestinal colonization by T cells carrying y8and afi T-cell antigen receptors were studied in nude and normal mice by flow cytometry and immunohistology. Furthermore, 'y and ag? T-cell development was analyzed in lethally irradiated mice that were reconstituted by fetal liver precursors with otf without a thymus. Our results establish that a major subpopulation of y8 intestinal intraepithelial lymphocytes is produced from uncommitted precursors at extrathymic sites. This work further shows that a small pool of T cells carrying a; T-cell receptors can also differentiate extrathymically from CD37 fetal liver precursors but with rates of production and peripheral expansion much reduced as compared with those observed in thymus-bearing animals.The thymus plays an essential role in the differentiation of T lymphocytes from precursors, as well as in the determination of mature T-cell repertoires. Thus, rearrangement of the receptor genes occurs in differentiating thymocytes, and antigens in the thymic environment shape T-cell repertoires (1)(2)(3)(4)(5). It has long been known that very few T cells are generated in athymic, nude animals (6, 7); these studies, however, analyzed peripheral lymphoid organs which harbor a very large excess of T lymphocytes bearing cra T-cell receptors (8,9). In contrast, recent observations have indicated that nude mice contain substantial numbers of y8 T cells (10, 11). The possible thymus-independent origin of y6-subset cells could contribute to our understanding of the specific characteristics, of this T-lymphocyte population, such as their selective localization in epithelia (12-17), the preferential variable region of T-cell receptor y-or 8-chain gene utilization at different anatomic sites (17-23), and the very limited repertoires in some epithelia (17,18,22,23 Cell Preparations. Spleens and mesenteric lymph nodes were removed in balanced salt solution without phenol red/ 3% fetal calf serum/0.015 M sodium azide (fluorescence medium). IEL cell suspensions were prepared as described (30) and according to standard procedures (31-33). In short, after being flushed extensively to eliminate the lumen content, small intestines were separated from Peyer's patches, longitudinally opened, and cut into pieces of 1-2 cm. Fragments were then. gently shaken in five changes of Ca2l and Mg2 -free Hanks' balanced salt solution and incubated with gentle stirring (100 rpm) for 7-10 min at room temperature in 25 ml of Ca2+-and Mg2+-free Hanks' solution/5 mM EDTA/ dithiothreitol at 70.ug/ml. Fragments were allowed to sediment for 5 min on ice after which the supernatant, containing cells, was recovered. IEL cells were washed three times and passed through nylon mesh and siliconized glass wool columns to remove cell aggregates and dead cells. No further purification step., such as Percoll gradients, was used. At this stage cell viability was scored in trypan blue, and fluorescence stainings were done.Immunofluorescence and Flowcytometric Analysis. Immunofluorescence st...

Regulatory T cells (T reg cells) constitute a population of CD4+ T cells that limits immune responses. The transcription factor Foxp3 is important for determining the development and function of T reg cells; however, the molecular mechanisms that trigger and maintain its expression remain incompletely understood. In this study, we show that mice deficient for the Ets-1 transcription factor (Ets-1−/−) developed T cell–mediated splenomegaly and systemic autoimmunity that can be blocked by functional wild-type T reg cells. Spleens of Ets-1−/− mice contained mostly activated T cells, including Th2-polarized CD4+ cells and had reduced percentages of T reg cells. Splenic and thymic Ets-1−/− T reg cells expressed low levels of Foxp3 and displayed the CD103 marker that characterizes antigen-experienced T reg cells. Thymic development of Ets-1−/− T reg cells appeared intrinsically altered as Foxp3-expressing cells differentiate poorly in mixed fetal liver reconstituted chimera and fetal thymic organ culture. Ets-1−/− T reg cells showed decreased in vitro suppression activity and did not protect Rag2−/− hosts from naive T cell–induced inflammatory bowel disease. Furthermore, in T reg cells, Ets-1 interacted with the Foxp3 intronic enhancer and was required for demethylation of this regulatory sequence. These data demonstrate that Ets-1 is required for the development of natural T reg cells and suggest a role for this transcription factor in the regulation of Foxp3 expression.

The generation of T cells depends on the migration of hematopoietic progenitor cells to the thymus throughout life. The identity of the thymus-settling progenitor cells has been a matter of considerable debate. Here we found that thymopoiesis was initiated by a first wave of T cell lineage-restricted progenitor cells with limited capacity for population expansion but accelerated differentiation into mature T cells. They gave rise to αβ and γδ T cells that constituted Vγ3(+) dendritic epithelial T cells. Thymopoiesis was subsequently maintained by less-differentiated progenitor cells that retained the potential to develop into B cells and myeloid cells. In that second wave, which started before birth, progenitor cells had high proliferative capacity but delayed differentiation capacity and no longer gave rise to embryonic γδ T cells. Our work reconciles conflicting hypotheses on the nature of thymus-settling progenitor cells.